SYNTHETIC OLIGODEOXYNUCLEOTIDE PURIFICATION VIA CATCHING BY POLYMERIZATION

Large quantities of pure synthetic oligodeoxynucleotides (ODNs) are important for preclinical research, drug development, and biological studies. These ODNs are synthesized on an automated synthesizer. It is inevitable that the crude ODN product contains failure sequences which are not easily removed because they have the same properties as the full length ODNs. Current ODN purification methods such as polyacrylamide gel electrophoresis (PAGE), reversed-phase high performance liquid chromatography (RP HPLC), anion exchange HPLC, and affinity purification can remove those impurities. However, they are not suitable for large scale purification due to the expensive aspects associated with instrumentation, solvent demand, and high labor costs. To solve these problems, two non-chromatographic ODN purification methods have been developed. In the first method, the full-length ODN was tagged with the phosphoramidite containing a methacrylamide group and a cleavable linker while the failure sequences were not. The full-length ODN was incorporated into a polymer through radical acrylamide polymerization whereas failure sequences and other impurities were removed by washing. Pure full-length ODN was obtained by cleaving it from the polymer. In the second method, the failure sequences were capped by a methacrylated phosphoramidite in each synthetic cycle. During purification, the failure sequences were separated from the full-length ODN by radical acrylamide polymerization. The full-length ODN was obtained via water extraction. For both methods, excellent purification yields were achieved and the purity of ODNs was very satisfactory. Thus, this new technology is expected to be beneficial for large scale ODN purification.

DEVELOPMENT OF CONFOCAL IMAGING TECHNIQUES FOR PROBING INTERFACIAL DYNAMICS IN MICROSCALE, GAS-LIQUID, TWO-PHASE FLOW

Micro-scale, two-phase flow is found in a variety of devices such as Lab-on-a-chip, bio-chips, micro-heat exchangers, and fuel cells. Knowledge of the fluid behavior near the dynamic gas-liquid interface is required for developing accurate predictive models. Light is distorted near a curved gas-liquid interface preventing accurate measurement of interfacial shape and internal liquid velocities. This research focused on the development of experimental methods designed to isolate and probe dynamic liquid films and measure velocity fields near a moving gas-liquid interface. A high-speed, reflectance, swept-field confocal (RSFC) imaging system was developed for imaging near curved surfaces. Experimental studies of dynamic gas-liquid interface of micro-scale, two-phase flow were conducted in three phases. Dynamic liquid film thicknesses of segmented, two-phase flow were measured using the RSFC and compared to a classic film thickness deposition model. Flow fields near a steadily moving meniscus were measured using RSFC and particle tracking velocimetry. The RSFC provided high speed imaging near the menisci without distortion caused the gas-liquid interface. Finally, interfacial morphology for internal two-phase flow and droplet evaporation were measured using interferograms produced by the RSFC imaging technique. Each technique can be used independently or simultaneously when.